Analysis of apoptosis levels in normal, inflamed and neoplastic colonic mucosa : the effects of non-steroidal anti-inflammatory drugs and 5-aminosalicylic acid on human colonic epithelial cell viability

Numerous epidemiological, in vivo and in vitro studies have demonstrated the chemopreventive potential of NSAIDs, and it is thought that this chemoprevention may be due to the induction of apoptosis within malignant cells. The 5-aminosalicyclic acid (5-ASA) compounds are structurally similar to the NSAID aspirin, and have also been shown to reduce tumour growth in recent studies. There are a number of mechanisms by which this chemoprevention may occur, including inhibition of cyclooxygenase expression, induction of apoptosis through alterations in expression of the Bcl-2 family members, and activation of the peroxisome proliferator activated receptor family of nuclear hormone receptors. The studies presented in this thesis were designed firstly, to analyse cell turnover in normal, inflamed and neoplastic colonic mucosa, and secondly to compare and contrast the effects of the NSAID indomethacin, and 5-ASA on human colonic adenocarinoma cell lines, and to gain further information regarding the mechanistic pathways underlying the chemopreventive properties of these compounds. Cell turnover was analysed in colonic mucosa using immunohistochemistry. The effects of indomethacin or 5-ASA treatment on cell viability, apoptosis and cell cycle distribution in human colonic cell lines was analysed by immunohistochemistry and flow cytometry. Changes in Bax, Bcl-XL and PPAR expression were analysed by RT-PCR. Indomethacin reduced cell viability, increased apoptosis, and caused cell cycle accumulation in the G0/G1 phase of the cell cycle. These effects were not dependant on cyclooxygenase expression. No changes to either cell viability or apoptosis levels were seen in cell lines following 5-ASA treatment. PPARgamma, and both PPARalpha and gamma expression increased following treatment of cell lines with 5-ASA or indomethacin respectively. In conclusion, these studies suggest that 5-ASA and indomethacin may have a different mode of action with regards to chemoprevention. Both 5-ASA and indomethacin may result in activation of the PPAR nuclear receptors, this presents an alternative mechanistic pathway for the observed chemopreventive properties of these compounds.